Project Summary/Abstract A better understanding of immune signaling mechanisms triggered by various types of antigens is critical for development of more effective vaccines against current and emerging pathogens. Our long-term goal is to understand how B cells integrate signals from antigens and their environment to produce effective immunity against pathogens while maintaining tolerance to self-derived antigens. The objective in this application is to determine the mechanisms by which a family of adhesion molecules, av integrins and autophagy proteins regulate germinal center (GC) B cell processing of viral antigens and how that impacts development of effective immunity to multiple Influenza virus strains. In published work we have shown that avb3 heterodimer from the av family, combines with components of the autophagy pathway, to limit TLR signaling in B cells. In current studies we find that, av-autophagy pathway limits GC B cells response to viral antigens containing TLR ligands. As a result, av-CD19 mice lacking av on B cells show increase in key features of GC-mediated antibody response such as affinity maturation of antibodies, generation of memory B cells and long-lived plasma cells, upon immunization with virus like particles or Influenza virus. Moreover, these mice develop increased cross-reactive antibodies against influenza virus strains after immunization with one strain. av-CD19 mice also develop increased autoantibodies with age, and we propose that av-mediated regulation of TLR signaling is a mechanism that limits excessive B cell responses to self-antigens. Our central hypothesis for this grant, is that the av-autophagy pathway also regulates endosomal processing of antigens containing TLR ligands, derived from pathogens, and limits GC B cell activation by these antigens. Therefore, removal of this control from GC B cells, enhances GC B cell TLR signaling and GC-mediated generation of antibodies with increased affinity and breadth against pathogens. Our rationale for this proposal is that a better understanding of new pathways regulating GC B cell activation by viral antigens will allow us to develop much needed vaccination strategies for generation of effective immunity against rapidly evolving viruses such as Influenza. In this grant we propose to: (1) Determine exactly how autophagy proteins are involved in GC B cell response to viral antigens; and (2) Determine whether removal of av-autophagy pathway can lead to better cross protective immunity to influenza virus strains; and (3) Determine how pathogen derived antigens containing TLR ligands get processed by GC B cells and how av-autophagy pathway alter this process. The proposed work is significant because it addresses the mechanisms of GC B cell activation by pathogen derived antigens, which is relatively unexplored but essential to understand for developing better vaccines. Moreover, based on these studies, av antagonists could be used to design vaccine adjuvants that provide long- term protection against wide variety of Influenza virus strains. Our approach is innovative as we are using a unique mouse model of enhanced B cell TLR signaling to investigate a novel TLR-av-autophagy pathway.